Ana Koren1, Eva Sodja2, Matija Rijavec2, Mojca Jez3, Viljem Kovac4, Peter Korosec2, Tanja Cufer2. 1. University Clinic Golnik, Golnik 36, 4204, Golnik, Slovenia. ana.koren@klinika-golnik.si. 2. University Clinic Golnik, Golnik 36, 4204, Golnik, Slovenia. 3. Blood Transfusion Centre of Slovenia, Slajmerjeva 6, 1000, Ljubljana, Slovenia. 4. Institute of Oncology Ljubljana, Zaloska cesta 2, 1000, Ljubljana, Slovenia.
Abstract
BACKGROUND: The rarity of circulating tumour cells (CTCs) in peripheral blood requires the application of sensitive techniques for their detection. The aim of our study was to (i) first determine the sensitivity of cytokeratin-7 (KRT7) mRNA expression levels for the molecular detection of CTCs using spiked-in lung adenocarcinoma (AC)-derived A549 cells and (ii) evaluate the impact of KRT7 mRNA expression in peripheral whole blood on the response to treatment and prognosis of patients with advanced lung AC who were treated with first-line platinum-based chemotherapy. METHODS: A549 cells were micro-manipulated before being spiked into whole blood samples obtained from healthy donors. Additionally, whole blood samples from 65 lung AC patients were collected in PAXgene blood tubes before the start of first-line platinum-based chemotherapy. KRT7 mRNA expression was measured using RT-qPCR. RESULTS: Through the spike-in experiment we found that it is feasible to detect a single A549 tumour cell in 2.5 ml whole blood and that the KRT7 mRNA levels were linearly correlated with the number of spiked-in tumour cells with a high reproducibility. In lung AC patients, no significant differences in response rate to chemotherapy, progression-free survival or overall survival and KRT7 mRNA levels were found. CONCLUSIONS: Our data show that KRT7 mRNA expression measured by RT-qPCR serves as a sensitive approach for the molecular detection of KRT7-positive CTC-resembling A549 cells in peripheral whole blood. The KRT7 mRNA levels measured were not significantly associated with the response to chemotherapy or the survival of patients with advanced lung AC. Additional studies are required to establish the possible clinical significance of KRT7 mRNA expression in whole blood after chemotherapy.
BACKGROUND: The rarity of circulating tumour cells (CTCs) in peripheral blood requires the application of sensitive techniques for their detection. The aim of our study was to (i) first determine the sensitivity of cytokeratin-7 (KRT7) mRNA expression levels for the molecular detection of CTCs using spiked-in lung adenocarcinoma (AC)-derived A549 cells and (ii) evaluate the impact of KRT7 mRNA expression in peripheral whole blood on the response to treatment and prognosis of patients with advanced lung AC who were treated with first-line platinum-based chemotherapy. METHODS: A549 cells were micro-manipulated before being spiked into whole blood samples obtained from healthy donors. Additionally, whole blood samples from 65 lung AC patients were collected in PAXgene blood tubes before the start of first-line platinum-based chemotherapy. KRT7 mRNA expression was measured using RT-qPCR. RESULTS: Through the spike-in experiment we found that it is feasible to detect a single A549 tumour cell in 2.5 ml whole blood and that the KRT7 mRNA levels were linearly correlated with the number of spiked-in tumour cells with a high reproducibility. In lung AC patients, no significant differences in response rate to chemotherapy, progression-free survival or overall survival and KRT7 mRNA levels were found. CONCLUSIONS: Our data show that KRT7 mRNA expression measured by RT-qPCR serves as a sensitive approach for the molecular detection of KRT7-positive CTC-resembling A549 cells in peripheral whole blood. The KRT7 mRNA levels measured were not significantly associated with the response to chemotherapy or the survival of patients with advanced lung AC. Additional studies are required to establish the possible clinical significance of KRT7 mRNA expression in whole blood after chemotherapy.
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